A comprehensive metatranscriptome analysis pipeline and its validation using human small intestine microbiota datasets.
A comprehensive metatranscriptome analysis pipeline and its validation using human small intestine microbiota datasets.
复制标题
DOI：
10.1186/1471-2164-14-530
复制
发表时间：
2013-08-02
期刊：
影响因子：
4.4
通讯作者：
Kleerebezem M
中科院分区：
文献类型：
--
作者：
Leimena MM;Ramiro-Garcia J;Davids M;van den Bogert B;Smidt H;Smid EJ;Boekhorst J;Zoetendal EG;Schaap PJ;Kleerebezem M
下一代测序 (NGS) 技术可通过直接 cDNA 测序（称为 RNA 测序 (RNA-seq)）应用于复杂的微生物生态系统中进行元转录组分析。 RNA-seq 生成非常复杂的大型数据集，对其进行全面解释需要可靠的生物信息学管道。在这项研究中，我们专注于开发这样一个元转录组管道，我们使用源自两个回肠造口个体的小肠微生物群的 Illumina RNA-seq 数据集对其进行验证。这里开发的元转录组管道能够有效去除 rRNA 衍生序列，然后对 mRNA 读数的预测功能和分类起源进行可靠的分配。小肠宏转录组数据集的系统发育分析显示，其与通过 16S rDNA 和 rRNA 焦磷酸测序获得的群落组成概况有很强的相似性，表明群落组成 (rDNA) 及其微生物成员之间总体 (rRNA) 和特异性 (mRNA) 活性的分类分布之间存在相当大的一致性。宏转录组测序方法的重现性是通过独立的重复实验建立的。此外，使用单端或双端测序方法进行宏转录组分析的比较表明，后一种方法不能提供改进的功能或系统发育见解。宏转录组功能图谱可以分析微生物群的全球和属特定活性，并说明这些方法在揭示微生物生态系统中互养相互作用的潜力。使用从人类小肠微生物群获得的 RNA-seq 数据集开发和评估了可靠的元转录组数据分析流程。管道的设置非常通用，可以应用于任何选定的利基中的（细菌）宏转录组分析。
Next generation sequencing (NGS) technologies can be applied in complex microbial ecosystems for metatranscriptome analysis by employing direct cDNA sequencing, which is known as RNA sequencing (RNA-seq). RNA-seq generates large datasets of great complexity, the comprehensive interpretation of which requires a reliable bioinformatic pipeline. In this study, we focus on the development of such a metatranscriptome pipeline, which we validate using Illumina RNA-seq datasets derived from the small intestine microbiota of two individuals with an ileostomy. The metatranscriptome pipeline developed here enabled effective removal of rRNA derived sequences, followed by confident assignment of the predicted function and taxonomic origin of the mRNA reads. Phylogenetic analysis of the small intestine metatranscriptome datasets revealed a strong similarity with the community composition profiles obtained from 16S rDNA and rRNA pyrosequencing, indicating considerable congruency between community composition (rDNA), and the taxonomic distribution of overall (rRNA) and specific (mRNA) activity among its microbial members. Reproducibility of the metatranscriptome sequencing approach was established by independent duplicate experiments. In addition, comparison of metatranscriptome analysis employing single- or paired-end sequencing methods indicated that the latter approach does not provide improved functional or phylogenetic insights. Metatranscriptome functional-mapping allowed the analysis of global, and genus specific activity of the microbiota, and illustrated the potential of these approaches to unravel syntrophic interactions in microbial ecosystems. A reliable pipeline for metatransciptome data analysis was developed and evaluated using RNA-seq datasets obtained for the human small intestine microbiota. The set-up of the pipeline is very generic and can be applied for (bacterial) metatranscriptome analysis in any chosen niche.
登录
查看更多内容
影响因子：
12.3
作者：
Giannoukos G;Ciulla DM;Huang K;Haas BJ;Izard J;Levin JZ;Livny J;Earl AM;Gevers D;Ward DV;Nusbaum C;Birren BW;Gnirke A
通讯作者：
Gnirke A
影响因子：
14.9
作者：
Cole JR;Wang Q;Cardenas E;Fish J;Chai B;Farris RJ;Kulam-Syed-Mohideen AS;McGarrell DM;Marsh T;Garrity GM;Tiedje JM
通讯作者：
Tiedje JM
影响因子：
48
作者：
Mortazavi, Ali;Williams, Brian A.;Wold, Barbara
通讯作者：
Wold, Barbara
影响因子：
2.3
作者：
Maccaferri, Simone;Biagi, Elena;Brigidi, Patrizia
通讯作者：
Brigidi, Patrizia
影响因子：
4.4
作者：
Klappenbach, JA;Dunbar, JM;Schmidt, TM
通讯作者：
Schmidt, TM